Goldman Visual Field (GVF) of the patient IV-4 at age 38 and at age 55.

<p>GVF demonstrated progressive loss from retinal disease. At age 38, both eyes showed a central “tunnel” vision of 15 degrees and large peripheral temporal islands OU (panel A and B), but by age 55, these had constricted to less than 5 degrees, with a small temporal islands for both eyes (panel C and D).</p

Levels of serum retinol in siblings with the RBP4 c.111+1 G>A change.

<p>Levels of serum retinol in siblings with the RBP4 c.111+1 G>A change.</p

Exome Analysis Identified a Novel Mutation in the RBP4 Gene in a Consanguineous Pedigree with Retinal Dystrophy and Developmental Abnormalities

<div><p>Retinitis Pigmentosa (RP) is a common form of retinal degeneration characterized by photoreceptor degeneration and retinal pigment epithelium (RPE) atrophy causing loss of visual field and acuities. Exome sequencing identified a novel homozygous splice site variant (c.111+1G>A) in the gene encoding retinol binding protein 4 (RBP4). This change segregated with early onset, progressive, and severe autosomal recessive retinitis pigmentosa (arRP) in an eight member consanguineous pedigree of European ancestry. Additionally, one patient exhibited developmental abnormalities including patent ductus arteriosus and chorioretinal and iris colobomas. The second patient developed acne from young age and extending into the 5^th decade. Both patients had undetectable levels of RBP4 in the serum suggesting that this mutation led to either mRNA or protein instability resulting in a null phenotype. In addition, the patients exhibited severe vitamin A deficiency, and diminished serum retinol levels. Circulating transthyretin levels were normal. This study identifies the RBP4 splice site change as the cause of RP in this pedigree. The presence of developmental abnormalities and severe acne in patients with retinal degeneration may indicate the involvement of genes that regulate vitamin A absorption, transport and metabolism.</p> </div

Electroretinogram responses from patient IV-4 with photopic single flash and 30 Hz flicker response stimuli.

<p>Both panels show responses at younger age 38 years on top and at 55 years on bottom. The single flash cone responses at age 38 was reduced approximately 80% from normal and was no longer detectable 17 years later (Panel A). The cone flicker response was similarly reduced at age 38 and further diminished by age 55 (Panel B).</p

Serum RBP4 and transthyretin (TTR) levels in affected and unaffected family members.

<p>Serum RBP4 and TTR levels were examined by western blot analysis as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0050205#s2" target="_blank">Methods</a>. A) The patients, IV-2 and IV-4 had undetectable levels of wild type RBP4 in contrast to their unaffected sibling and the control. A faint band was observed in the region of predicted mutant RBP4 in both patients and their unaffected sibling (Asterisk). B) TTR levels (TTR monomer (15 kDa) and dimer (30 kDa) were indistinguishable between affected and unaffected subjects.</p

Genetic analysis of a family with retinal degeneration.

<p>A. Pedigree of the family with retinal degeneration. Family members that were available for study were genotyped for the RBP4 c.111+1 G>A mutation. B. The reverse complement sequence of the heterozygous subject IV-3 (G/A) and the homozygous subject IV-2 (A/A) are shown. C. Protein encoding transcripts of <i>RBP4</i> reported by <i>Ensembl</i> (<a href="http://uswest.ensembl.org" target="_blank">http://uswest.ensembl.org</a>). Three protein coding transcripts are reported by <i>Ensembl</i>. Transcripts ENST00000371464 and ENST00000371467 encode the same protein (CCDS 31249). ENST00000371469 does not have a CCDS identifier. The <i>RBP4</i> c.111+1 mutation would affect all three of the predicted protein coding transcripts.</p

Investigating the Molecular Basis of Retinal Degeneration in a Familial Cohort of Pakistani Decent by Exome Sequencing

<div><p>Purpose</p><p>To define the molecular basis of retinal degeneration in consanguineous Pakistani pedigrees with early onset retinal degeneration.</p><p>Methods</p><p>A cohort of 277 individuals representing 26 pedigrees from the Punjab province of Pakistan was analyzed. Exomes were captured with commercial kits and sequenced on an Illumina HiSeq 2500. Candidate variants were identified using standard tools and analyzed using exomeSuite to detect all potentially pathogenic changes in genes implicated in retinal degeneration. Segregation analysis was performed by dideoxy sequencing and novel variants were additionally investigated for their presence in ethnicity-matched controls.</p><p>Results</p><p>We identified a total of nine causal mutations, including six novel variants in <i>RPE65</i>, <i>LCA5</i>, <i>USH2A</i>, <i>CNGB1</i>, <i>FAM161A</i>, <i>CERKL</i> and <i>GUCY2D</i> as the underlying cause of inherited retinal degenerations in 13 of 26 pedigrees. In addition to the causal variants, a total of 200 variants each observed in five or more unrelated pedigrees investigated in this study that were absent from the dbSNP, HapMap, 1000 Genomes, NHLBI ESP6500, and ExAC databases were identified, suggesting that they are common in, and unique to the Pakistani population.</p><p>Conclusions</p><p>We identified causal mutations associated with retinal degeneration in nearly half of the pedigrees investigated in this study through next generation whole exome sequencing. All novel variants detected in this study through exome sequencing have been cataloged providing a reference database of variants common in, and unique to the Pakistani population.</p></div

Members of the pedigree enrolled in this study are indicated by the presence of a genotype below their icon.

<p>“+” indicates presence of the variant, “-” indicates absence of RPE65 variant (NM_000329.2:c.1087 C>A, NP_000320.1:p.Pro363Thr). Icons representing the individual(s) selected for exome capture and sequencing are outlined in red. A question mark (?) within the icon indicates the patient was not clinically examined. (A) PKRD281. (B) PKRD282. (C) PKRD283. (D) PKRD284. (E) PKRD285.</p

Members of the pedigree enrolled in this study are indicated by the presence of a genotype below their icon.

<p>“+” indicates presence of the variant, “-” indicates absence of the variant indicated to the left of the pedigree. Icons representing the individual(s) selected for exome capture and sequencing are outlined in red. A question mark (?) within the icon indicates the patient was not available for clinical evaluation. (A) PKRD077. (B) PKRD078. (C) PKRD103. (D) PKRD138. (E) PKRD141. (F) PKRD142. (G) PKRD176 (H) PKRD185.</p

Electroretinography recordings of affected individuals with inherited retinal degeneration.

<p>Scotopic 0 dB response, and photopic 0 dB 30Hz flicker response of A) OD and B) OS of individual VI:9 of family PKRD138; C) OD and D) OS of individual V:5 of family PKRD141; E) OD and F) OS of individual VI:2 of family PKRD142; G) OD and H) 374 OS of individual V:7 of family PKRD176; I) OD and J) OS of an unaffected control. The electroretinography responses of affected individuals illustrate loss of rod and cone response. OD = oculus dexter (right eye); OS = oculus sinister (left eye).</p